Industrial fermentations predominately use glucose as a feed stock for the production of a multitude of proteins, enzymes, alcohols and other chemicals. In many applications, the glucose is produced by the enzymatic conversion of starch. This conversion is frequently accomplished by a two-step process. The first step is a liquefaction step, wherein an insoluble granular starch substrate is slurried in water, gelatinized with heat and hydrolyzed by a thermostable alpha amylase (e.g., E.C. 3.2.1.1: 1,4-alpha-D-glucan glucoanohydrolase) in the presence of calcium. The second step is a saccharification step, wherein the soluble dextrins (sugars) produced in the first step are further hydrolyzed to glucose by an enzyme having glucoamylase (e.g., E.C. 3.2.1.3: 1,4-alpha-D-glucan glucohydrolase) activity. Glucoamylases catalyze the release of glucose from the non-reducing ends of starch. Glucose may then be used as an end product or used as a precursor to be converted to other commercially important end products, such as fructose, ethanol, ascorbic acid (ASA) intermediates and/or 1,3-propanediol.
Therefore, glucoamylases, which are involved in the conversion of starch to sugar are extremely important industrial enzymes. Glucoamylases may be obtained from bacteria, plants and fungi. However, preferred glucoamylases are derived from fungal strains. Examples of fungal glucoamylases include those obtained from strains of Aspergillus, Rhizopus, Humicola and Mucor (See, WO 92/00381 and WO 00/04136).
Various glucoamylases have been commercialized, including Aspergillus nigerglucoamylase (e.g., trade name OPTIDEX L-400® from Genencor International Inc. and trade name AMG from Novo Nordisk) and Rhizopus (e.g., trade name CU.CONC from Shin Nihon Chemicals, Japan and trade name GLUCZYME from Amano Pharmaceuticals, Japan).
Certain thermophilic and mesophilic fungi, and particularly strains of Humicola grisea and Aspergillus awamori, produce an enzyme having both glucoamylase activity and the ability to hydrolyze raw starch. These glucoamylases are referred to as granular starch hydrolyzing enzymes (GSHE) and are also known in the art as raw starch hydrolyzing (RSH) enzymes. Additionally, while these enzymes will hydrolyze thinned starch hydrolyzate to glucose in a manner similar to other known glucoamylases, they frequently have a pH optimum in the range of 5.0 to 7.0 as compared to a pH optimum of less than 5.0 for widely used glucoamylase preparations (See, Tosi et al., (1993) Can. J. Microbiol., 39: 846-851).